1. Technical Field of the Invention
The present invention relates generally to the fields of microbiology, infectious diseases and drug toxicity. More specifically, the present invention relates to methods of modeling infectious disease and chemosensitivity with three-dimensional suspension of mammalian cells using a bioreactor.
2. Description of the Prior Art
An essential feature of the pathogenicity of microorganisms is their ability to infect or colonize host epithelial cells and cells in other tissues. The terms “infect” and “colonize” are used interchangeably herein, and encompass adherence to cells, invasion, survival within, and damage or destruction of the cells. Initial sites of bacterial infection include the epithelia of the intestine, kidney, lung and bladder. There is a need to investigate how bacterial and other pathogens interact with the epithelium to initiate disease, which can be through the use of both in vitro and in vivo models of infection.
Some types of tissue culture models of infectivity are not the best models of the conditions faced in vivo by pathogens. Conclusions drawn about the interaction between pathogens and the host epithelium using cultured epithelial cells may be limited due to the dedifferentiation of these cells during conventional cell culture. Many of the physiological differences between cultured cells and their in vivo counterparts are believed to be the result of the dissociation of cells from their native three-dimensional geometry in vivo to their propagation on a two-dimensional substrate in vitro. Likewise, many of the characteristics of animal models fail to mimic the human disease, and animal models present a complex system in which many variables cannot be controlled. Accordingly, since certain in vitro nor in vivo assays do not replicate the complex environment encountered by pathogens during the natural course of human infection, the information gained from these studies may be limited.
An essential feature of the pathogenicity of Salmonella is its ability to invade host intestinal epithelial cells. While important advances have been made toward understanding how Salmonella interacts with the intestinal epithelium to initiate disease, through the use of both in vitro and in vivo models of infection, numerous questions have been derived from these studies which remain to be answered. In particular, it is well-documented that important differences exist between the pathogenesis of serovar Typhimurium in human infections and in widely-used cell culture and animal models. Specifically, tissue culture models of infectivity are not exact models of the conditions faced in vivo by Salmonella. Conclusions drawn about the interaction between Salmonella and the host intestinal epithelium using cultured enterocytes are limited due to the dedifferentiation of these cells during conventional cell culture. Many of the physiological differences between cultured cells and their in vivo counterparts are believed to be the result of the dissociation of cells from their native three-dimensional geometry in vivo to their propagation on a two-dimensional substrate in: vitro. Likewise, many characteristics of animal models fail to mimic the human disease, and animal models present a complex system in which many variables cannot be controlled. Accordingly, since neither in vitro nor ill vivo assays faithfully replicate the complex environment encountered by Salmonella during the natural course of human infection, the information gained from these studies is limited. A high fidelity enteric cell culture model can provide new insights into studies of Salmonella infectivity by bridging the gap between the inherent limitations of cultured mammalian cells and intact animals.
The response of cells to various drugs is an area of great interest to the medical field. Currently, many model systems exist for testing the chemosensitivity of cells to various chemical compounds in order to identify new pharmaceutical agents for the treatment of cancer and other diseases. Conventional cell culturing techniques do not always accurately simulate conditions ill vivo favorable for the growth of some tissues. Hence, some biological properties affecting tissue response to drugs are not apparent with standard cell culturing methods.
U.S. Pat. No. 6,117,674, which is fully incorporated by reference herein, discloses an invention relating to the propagation of a pathogen selected from the group consisting of viruses, bacteria, protozoans, parasites and fungi, by inoculating a three dimensional tissue mass culture at microgravity conditions in fluid culture media in a microgravity vessel with the pathogen. Specifically, the microgravity conditions are simulated in unit gravity by a horizontal rotating wall vessel (RWV). Growth conditions in the RWV allow for better cellular differentiation and formation of three-dimensional cellular aggregates, more efficient cell-to-cell interactions, the ill vivo-like exchange of growth factors and greater molecular scaffolding facilitating mechanical stability for cells. The RWV bioreactors offer a revolutionary approach not previously applied for studying microbial infectivity from the perspective of the host-pathogen interaction and also for analyzing chemosensitivity to toxins and chemotherapeutic agents, like antibiotics.